肾小管上皮细胞C/EBPα条件性敲除小鼠的构建及鉴定  

Generation and identification of renal tubular epithelial cell C/EBPα conditional knockout mice

作　　者：应霁 刘剑[1] 张沥文[1] 仲芳[1] 周芳芳[1] 王伟铭[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院肾脏科,上海200025

出　　处：《上海交通大学学报（医学版）》2016年第10期1403-1407,共5页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金(81270782;30771000);国家重点基础研究发展计划("973"计划)(2012CB517701;2012CB517604);科技部十二五科技攻关项目(2011BAI10B00;2011BAI10B06);上海市卫计委重中之重学科建设项目;卫生行业科研专项项目(201002010);比较性研究;上海市科研计划项目(15140902800)~~

摘　　要：目的·构建肾小管上皮细胞C/EBPα条件性敲除小鼠,并进行表型鉴定,为进一步研究C/EBPα在肾间质纤维化中的作用奠定基础。方法·利用C/EBPα^(flox/flox)和pepck-Cre^+转基因小鼠进行饲养和杂交;将繁殖产生的第1代小鼠再次进行交配,得到第2代小鼠。通过PCR鉴定出基因型为C/EBPα^(flox/flox).pepck-Cre^+的小鼠(C/EBPα^(-/-)小鼠)。采用RT-q PCR及免疫组织化学法分别检测肾组织中C/EBPαmRNA和蛋白表达水平。应用H-E染色、Masson染色观察6~8周龄C/EBPα^(-/-)小鼠及野生型(WT)小鼠肾脏组织形态学改变。结果·2种转基因小鼠繁殖产生的第2代小鼠基因型符合孟德尔遗传定律,交配后获得C/EBPα^(-/-)小鼠。与WT小鼠相比,C/EBPα^(-/-)小鼠肾脏中C/EBPαmRNA及蛋白表达水平下降,光学显微镜下肾脏组织形态无明显改变。结论·应用Cre-loxp系统可以成功地敲除小鼠肾小管上皮细胞中C/EBPα基因,且在基础情况下C/EBPα^(-/-)小鼠肾脏组织无明显形态学改变。Objective·To generate renal tubular epithelial cell C/EBPα conditional knockout mice, perform phenotypic identification, and lay a foundation for further study on the role of C/EBPα in renal interstitial fibrosis.Methods·C/EBPα^flox/floxand pepck-Cre^＋ transgenic mice were bred and hybridized.The first generation offspring were mated to reproduce the second generation. Mice with C/EBPα^flox/floxpepck-Cre^＋ genotype （C/EBPα^-/-mice） were identified using PCR. The mRNA and protein expressions of C/EBPα in renal tissues were measured using RT-qPCR and immunohistochemistry staining. The morphologic changes in renal tissues of C/EBPα^-/-mice and wide type （WT） mice aged 6-8 weeks were observed using H-E staining and Masson staining.Results·The genotype of the second generation offspring of two types of transgenic mice accorded with the Mendelian laws and C/EBPα^-/-mice were generated. C/EBPα^-/-mice had lower mRNA and protein expressions of C/EBPα in renal tissues and no significant renal morphologic changes compared with WT mice.Conclusion·Cre-loxp system can successfully knock out the C/EBPα gene in mouse renal tubular epithelial cells. C/EBPα^-/-mice have no significant renal morphologic changes in baseline condition.

关 键 词：Cre-loxp系统 C/EBPΑ 条件性敲除 肾小管上皮细胞 

分 类 号：R692[医药卫生—泌尿科学]